Whether ionizing radiation can induce mammalian mutations by intragenic (or "point") mutation is not known with certainty. Evidence from studies using animals and cells indicates that most or all ionizing radiation mutagenesis is due to chromosomal aberration and not intragenic mutation. The hypothesis that it is possible for ionizing radiation to cause intragenic mutation will be tested using cultured human fibroblasts, lymphocytes, and epithelioid cells and ouabain resistance as biochemical marker. The induction of ouabain resistance is thought to be through a mutation within the mammalian gene for Na+K+ATPase, the membrane ion-transport protein afftected by this drug. Research to date, based on the post-irradiation selection of cells that can survive ouabain treatment and form colonies, has shown no evidence that this mutation can be caused by ionizing radiation. The proposed research will therefore seek ouabain-resistant mutant cells by a more sensitive and less survival-dependent technique: laser-activiated cell sorting. The effect of ouabain on gamma-irradiated and control cells will be measured using fluorescent dyes that measure membrane potential, and unaffected cells will be sorted and analysed for acquired and heritable ouabain resistance. If an increased number of such cells occurs in gamma-irradiated cultures an estimate of intragenic mutaton inducation rate will be available for the first time.